A study will be made of the active site of and the mechanism of hydrolysis by acetylcholinesterase; and ligands for the nicotinic cholinergic receptor will be devised, for analysis of the mechanism of permeabilty caused by the receptor. Much of the study derives from our view that the part of the active site of the enzyme which is generally considered to be an anionic subsite is not negatively charged, and may be termed the Trimethyl site, and that the active site may be more specifically explored by reagents containing t-butyl rather than trimethyl ammonium groups. Substrates for the enzyme will be prepared which will allow analysis, by kinetic study, of the volume and polar character of the Trimethyl site. Charged and uncharged reversible inhibitors for the enzyme, largely competitive, will be devised and applied in a detailed kinetic exploration of the nature of the parts of the active site. Parts of the enzyme active site will be covalently labelled by reactive compounds containing t-butyl groups - aziridinium, sulfonium, haloacetyl, active halogen, peroxy and diazo compounds. Binding of a positively charged group to an uncharged site may provide a mechanism for inducing conformational change, and this may occur on the receptor. Interaction of uncharged ligands with conformational forms of the receptor will be analyzed. Novel nicotinic non-competitive reversible and covalently bound antagonists will be devised for characterization of the local anaesthetic binding site and ion translocation channel.